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Zaia Draws on Decades of Innovation in Infectious Disease for Breakthroughs in Gene Therapy

Publication
Article
Evidence-Based OncologyJune 2020
Volume 26
Issue 5

Known as a gene therapy pioneer, Zaia has spent almost 40 years at City of Hope, in Duarte, California. He was first drawn by the promise of studying cytomegalovirus. Over the decades, his groundbreaking research has encompassed HIV/AIDS, cellular gene transfer therapy, immunotherapy, bispecific antibodies, and now hyperimmune globulin for workers on the frontlines of the coronavirus disease 2019 (COVID-19) pandemic.

https://doi.org/10.37765/ajmc.2020.43620How did discoveries in HIV research lead to the revolution of immuno-oncology? To understand this path, Evidence-Based Oncology™ spoke with John A. Zaia, MD, the Aaron D. Miller and Edith Miller Chair in Gene Therapy at City of Hope, a comprehensive cancer center. He also serves as director of its Center for Gene Therapy and is program director of the City of Hope Alpha Stem Cell Clinic, which is funded by the California Institute for Regenerative Medicine.

Known as a gene therapy pioneer, Zaia has spent almost 40 years at City of Hope, in Duarte, California. He was first drawn by the promise of studying cytomegalovirus. Over the decades, his groundbreaking research has encompassed HIV/AIDS, cellular gene transfer therapy, immunotherapy, bispecific antibodies, and now hyperimmune globulin for workers on the frontlines of the coronavirus disease 2019 (COVID-19) pandemic. Zaia was recently awarded $750,000 from the California Institute for Regenerative Medicine to study the potential use of convalescent plasma in patients with COVID-19, as well as to create the COVID-19 Coordination Program to aid in this effort.1

Zaia spoke at length about the crucial connections between basic research in HIV/AIDS and developments in gene therapy. This interview has been edited slightly for clarity.

EVIDENCE-BASED ONCOLOGY™ (EBO): We know that HIV does not elicit a protective immune response in the body. Do you think it is possible to overcome nature in this regard, to trick the immune system into fighting HIV to the degree that it can overcome it, such as interrupting the binding of the virus to the CD4 receptor?

ZAIA: So, let’s take that question apart. There is an immune response in the body to HIV, but it’s just not protective. The question is, why isn’t it protective? And could you overcome that deficiency? So, one aspect to understand is the ability of the virus to continuously mutate.

If you’re familiar with RNA replication, it doesn’t have high fidelity, meaning that mistakes occur while copying the new strand of RNA. Whereas DNA replication has high fidelity, meaning that once you copy it, it’s virtually word-for-word precise with only an occasional mutation. So, whenever the virus makes 10,000 base pair copies, there’s 1 mistake. But the virus is actually only 10,000 base pairs long in terms of its RNA. So that means there’s about 1 naturally occurring mistake in every new virus. And since there could be billions of new viruses made, there will be literally all these mutations a day, some of which could help the virus survive. Since the barriers that put up against the virus for continuing its replication are limited (eg, immune response, antiviral medications), it’s not that hard to imagine that a mutation could occur that gets around a specific barrier When this occurs, it is called “antigen escape” or “drug resistance.”

The deeper understanding of this question is, in the immune recognition of the virus, the T lymphocytes have a receptor for the virus called the T-cell receptor; it’s really an antigen receptor that can see a specific peptide on the surface of the virus or on the infected cell. So, the T-cell receptor itself is the problem. It’s exerting this selection, but it is not very flexible—it’s rigid. It’s an all-or-none thing. If the virus can mutate its protein slightly, then that peptide never fits into the receptor. It’s kind of like a lock and key. So, we need a T-cell receptor that’s more resistant to antigen escape. Could you make an artificial receptor, called a chimeric antigen receptor (CAR), that would better resist antigen escape? At City of Hope, we’ve been trying to make a T-cell receptor that you [could] paste on to the T cells genetically so they are better at resisting this inability to detect the mutated part of the virus. Is there a part of the virus that is resistant to mutation? There probably is. The key surface protein is called gp120. And some antibodies are very broadly reactive to all viruses, all HIV viruses. So, a broadly neutralizing antibody can detect multiple different gp120s, all of which are slightly different—but there’s some common feature that’s recognized by the broadly neutralizing antibodies. If you put that on a T cell, as a chimeric antigen receptor, the T cell might be more resistant to antigen escape by the mutating virus.

The other possibility is, what if you use the CD4 receptor? That’s an almost immutable part of the virus biology, because if the virus didn’t bind to the CD4 receptor, it probably wouldn’t be HIV. It would be a different virus, a different lentivirus. But there are CAR T-cell receptors that utilize not the antibody to find the virus, but the CD4 receptor itself to find the virus. In other words, if you put CD4 on the surface of a CD8 cell, it would find all the gp120 because the CD4 and gp120 would bind to each other. So that is actually another concept that can be utilized, and that’s currently in clinical trials at the University of Pennsylvania.2

So, in summary, I think the trick would be to utilize a modification of a T-cell receptor that would avoid the ability of the virus to mutate around the classical T-cell receptor and allow the immune system to see the virus and to control it.

EBO: The holy grail of HIV research for nearly 40 years has been to produce a vaccine. The Thai trial (RV144)3 has been the only trial thus far to show that a preventive HIV vaccine is possible. What have been the barriers to reproducing these results? Why has achieving the goal of developing an AIDS vaccine been so elusive?

ZAIA: Those are good questions. I don’t think anyone knows [the answers] for sure. But 2 factors are probably important. Again, you go back to the virus mutation issue. It’s continuous, and now [it’s] in the presence of immune pressure placed on the virus by the vaccine. You’ll get selection for these mutations. So, I guess the question is, did the vaccine make an immune response to the most immutable parts of the virus? Probably not—virus mutation is not the only answer to why vaccines fail. It seems to be something more basic than that.

For example, do vaccines induce mucosal immunity? We have a mucosal immunity to many viruses that come in contact with our mucous membranes via nose, throat, etc. Well, HIV would be in the mucous membranes of the genital tract and rectum, [usually]. So, are these vaccines really making a mucosal immunity where it’s needed? That’s a possible explanation for why the vaccines are not working.

An area that people just don’t understand at the present time is why you can make a vaccine for certain viruses that would normally come through the respiratory tract and not be able to make a vaccine for others. It’s relevant to COVID. Will we be able to make an immune response to COVID when we know that the initial entry point is through the nasal passages? That’ll be the million-dollar question.

EBO: Can you explain what a lentiviral vector is? Why is it that HIV can be inactivated outside the body to be used as a safe lentiviral gene therapy, but we can’t do the same with the virus internally?

ZAIA: A lentivirus has a certain structure and is made of RNA and protein. And it fulfills the requirements from some taxonomic committee that defines what a lentivirus is. Basically, it is a virus that can do 1 thing very usefully: it can reverse the RNA to DNA, and the DNA can then be integrated into the host DNA, become part of the host. And that integration is due to an enzyme called integrase. So, it has an RNA that also encodes for this integrase as well as reverse transcriptase; it turns RNA into DNA. That was a famous discovery at one point; it won the Nobel Prize. And so that’s what makes it a lentivirus.

Now, you can inactivate it in the sense of making it safe. It has only 9 major proteins, and those proteins are important in those elements that I just mentioned and in leading to its pathogenicity. You can remove them and still have some of the elements that you need. For example, you could leave the integrase but remove other things, which may make the virus able to replicate and lead to AIDS. But now you’d have an incomplete virus that you can put a gene into, and it can be delivered to the cell, because the virus can still get into the cell. And it can still have an integrase, which can help you integrate that message or that gene into the host cell. But the virus can’t replicate. It has all the other parts of it that are needed, but replication has been removed. You basically neuter the virus by removing critical genes. It’s still allowed to be a good virus for your use—it can get it into the cell, deliver its payload—but it just can’t replicate.

The question is, why can’t we inactivate certain of these critical genes that are important for replication? And, in fact, you can—in vitro. You can certainly put in inhibitors of all the various proteins of a virus. Some of those are called small inhibitory RNAs (siRNA), which are known to block specifically different proteins of a virus and almost any virus. The question is, how do you deliver that siRNA to all cells that are infected? If you have trillions of cells infected, how would you get to the last one? That’s the issue. So, you can do it in a test tube, but you just can’t do it in a human organism.

Now, you might ask, why are you able to get certain things to work for acute lymphoblastic leukemia but not for HIV?

Well, I think it’s because the virus can become latent and invisible to most systems and the leukemia cannot. The leukemia is robustly growing and expressing all of its proteins and enzymes, and so we [can fight it with] chemicals and other things like CAR T cells, and they will destroy those cells. The HIV is holed up in an inactive form, a so-called reservoir, and that reservoir is the problem. Once it’s in there, it’s like a snake in its hole. You cannot get to it.

EBO: In a 2016 commentary,4 you discuss the findings by Yang et al that the immune-mobilizing monoclonal T-cell receptor, or ImmTAV, could be an effective new agent against HIV/AIDS due to its bispecific antibody—binding properties. It was shown to have activity against both p17-expressing activated and resting CD4 cells. Is the agent proposed by Yang possible and without neurotoxicity?

ZAIA: Well, bispecific antibodies certainly work pretty well with cancer. We’ve actually made a bispecific [antibody] for HIV, where one end of it recognizes the envelope of the virus and the other end of it binds to a T cell, the CD8 T cell. And in vitro, that will suppress HIV. In fact, that work was presented at the virtual meeting of the American Society of Gene and Cell Therapy.

The question is, can you make a bispecific antibody easily? Sometimes it’s easy, and sometimes it’s not. One of the chemists here at City of Hope, Jack Shively, PhD, has worked out a method by which you can quickly bind 2 antibodies together so that they both maintain their specificity. If you think of the antibodies as having an active and an inactive end, you bind the 2 inactive ends together so that the active ends are sticking outward, and now you have whatever the specific activity was of each of the antibodies.

And so, we’ve taken a CD3 and joined it to gp120 antibody, and we’re able to suppress HIV. In theory, this is possible. Why hasn’t it been done? It’s one of those things where you’ve got to get the right motivation and the right company involved that has that information and the resources, and I think it will work. The limiting thing at this time, like any HIV cure research, is that most companies believe a pill a day is working pretty well for most people. Therefore, there is no significant market—whereas in cancer there is a greater need, and so, if one is going to make bispecific antibodies, why not make them for cancer and not risk the resources needed for an HIV product? If you could de-risk this process financially—i.e., the group came up with money to do a clinical trial for a company—I suspect that you would have some activity. At the present time, there is no such group.

EBO: Also in 2016, Johns Hopkins gained the first approval to perform organ transplants between HIV-positive individuals.5 By November of 2017, 34 such transplants had been performed at 6 hospitals. And in March 2019, the first kidney transplant was performed between living HIV-positive donors. In addition, you published results a decade ago showing that HIV-positive status should not preclude these individuals from transplant trials.6 Will HIV-positive living transplant donation move forward?

ZAIA: What we showed was that an HIV-positive person with lymphoma does just as well within an autotransplant setting as does anybody else who has a lymphoma. It wasn’t that we were saying that an HIV-positive person should become a donor for an HIV-negative person. It was purely in an autologous setting.

So, the real question is, will an HIV-positive living transplant donation move forward into an HIV-negative person? No, I don’t think that will happen unless someday persons living with HIV can become blood donors. And that’s an interesting area. For example, could they become plasma donors or even platelet donors? At the present time, they cannot. That’s the one area that is still hard to imagine: that the person living with HIV who has latent virus in his blood cells would ever be allowed to be a blood donor because those blood cells can then reactivate in the recipient and transfer the HIV. I think this question comes up a lot, especially with the question of COVID and of convalescent plasma. Plasma is the one place where it could first happen, as you can add inactivators to plasma that will inactivate viruses, but the American Association of Blood Banks and others in that community are strong advocates of a safe blood product. And in fact, every country is like that. They are very protective when it comes to blood products. And I don’t think in our lifetime a person living with HIV will ever become a blood donor for a person who does not have HIV.

EBO: In a 2011 interview, Anthony Fauci, MD, said there were aspects about HIV that were very unique and extremely frustrating—specifically, that it can’t be eradicated with drugs, although it can be suppressed well enough to give HIV-positive people a normal life. Fauci has since been tasked with creating a plan to eradicate HIV by 2030. Is this possible?

ZAIA: Well, the word eradicate is a critical question. We have eradicated smallpox from the planet, except for some freezers in which it still exists in the United States, Russia, and maybe some other places. But we have not eradicated polio from the planet. And yet we don’t think of polio as a problem, at least in Western countries, and it only exists in a few pockets now in Pakistan and other areas that don’t get adequate resources for managing disease like this that can be prevented. Do we want HIV to look like polio? Maybe. Maybe there will be small pockets of HIV. It may even eventually be like leukemia.

There’ll be an occasional case, not a whole lot. But maybe between now and 2030 there’ll be methods to treat patients who have HIV/AIDS with some of our developed gene therapies. But it won’t be truly eradicated from the planet like smallpox. I don’t think we can eradicate it completely. It’s possible you could take a pill so the virus is not being transmitted. And with time, let’s say you have 2 or 3 generations of people, one may imagine that there would be a smaller and smaller population that would have HIV, either couldn’t get the pill or, for some reason, the pill didn’t suppress their virus. So then, 2 or 3 generations later, the virus is still in small pockets. Something like that could happen, but I don’t see HIV being eradicated, although it could definitely be suppressed dramatically, as polio has been.

EBO: You are currently studying gene transfer for HIV-related therapy. How will this work potentially translate into clinical practice?

ZAIA: We know of people who have had bone marrow transplants with naturally genetic mutations—specifically, CCR5 mutation—in their stem cells. And we know that in that setting, it is possible to cure a patient of their HIV and have them exist without that viremia, that virus in their blood, without taking drugs. So that’s the idea. We would like to establish a gene, or a group of genes, that are placed in a stem cell and then transplanted into a patient. The problem, of course, is that we don’t have an optimal way to efficiently and safely transplant patients today. We have a busy transplantation patient program for leukemia and for life-threatening diseases, but those are fairly harsh therapies. Even now, we’re pleased when the failure rate is only 35% or 40%, meaning that almost two-thirds of the patients survived, because generally, with the transplantation, all are destined to die of their ailment.

We can’t apply these harsh methods yet in the setting of HIV to otherwise healthy people who are basically controlling their HIV on pills. But what if we could? So, that’s what’s needed. Right now, we don’t have a safe way to engraft the cells into a recipient, but it’s coming along. It may well happen that you could very safely administer stem cells. The next question is, when do they actually have an advantage over other stem cells in the body and help grow them and produce progeny. That would be when they’re protected. So that’s the other aspect: How do you get them to really compete well? We know how to do that, in general, but [for HIV,] it has never been specifically tested. And yet, it could happen. So, if those 2 things happen—a safe transplant and a way to make sure that the protected stem cells have an advantage—

then I think that gene transfer for HIV-related therapy will happen. We’ve completed a study of 8 patients in which we did genetically modify the cells, and we got the cells to engraft and we got them to persist but not at levels necessary to replace the infected cells. But this is a start, to which one can apply the metaphor of Goddard’s first liquid-fueled rocket that he set up in the 1930s in Worchester, Massachusetts. It was the first liquid-fueled rocket and it went up a brief distance; it might have gone up a mile and then fell to the earth, but it was the beginning of things. You knew that, in theory, it was possible. We’re not yet at the point where we know we can land on the moon yet in this area, but I think we know that it is possible and could happen in the next years. References

1. Bonar S. City of Hope to begin CIRM-funded coronavirus study. City of Hope. April 29, 2020. Accessed May 24, 2020. www.cityofhope.org/breakthroughs/cirm-funds-city-of-hope-coronavirus-study

2. Evaluating the safety and immunogenicity of env (A,B,C,A/E)/gag (C) DNA and gp120 (A,B,C,A/E) protein/GLA-SE HIV vaccines, given individually or co-administered, in healthy, HIV-1-uninfected adults. Updated November 26, 2019. Accessed May 24, 2020. https://clinicaltrials. gov/ct2/show/NCT03409276

3. Karasavvas N, Billings E, Rao M, et al. The Thai phase III HIV type 1 vaccine trial (RV144) regimen induces antibodies that target conserved regions within the V2 loop of gp120. AIDS Res Hum Retroviruses. 2012;28(11):1444-1457. doi:10.1089/aid.2012.0103

4. Zaia JA. A new agent in the strategy to cure AIDS. Mol Ther. 2016;24(11):1894-1896. doi:10.1038/mt.2016.194

5. One historic HIV organ transplant, numerous team members. News release. Johns Hopkins Medicine; February 21, 2017. Accessed May 24, 2020. hopkinsmedicine.org/news/articles/one-historic-hiv-organ-transplant-numerous-team-members

6. Krishnan A, Palmer JM, Zaia JA, et al. HIV status does not affect the outcome of autologous stem cell transplantation (ASCT) for non-Hodgkin lymphoma (NHL). Biol Blood Marrow Transplant. 2010;16(9):1302-1308. doi:10.1016/j.bbmt.2010.03.019

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